Chemical and biological investigations of Vietnamese cyanobacteria [Elektronische Ressource] / vorgeelgt von Le Thi Anh Tuyet

Ernst Moritz Arndt-Universität Greifswald
Mathematisch-Naturwissenschaftliche Fakultät




Chemical and Biological
Investigations of Vietnamese
Cyanobacteria

Inauguraldissertation
zur
Erlangung des akademischen Grades
doctor rerum naturalium (Dr. rer. nat.)
an der Mathematisch-Naturwissenschaftlichen Fakultät
der
Ernst-Moritz-Arndt-Universität Greifswald

vorgelegt von Le Thi Anh Tuyet
geboren am 19.05.1973
in Thanh Hoa, Vietnam







Greifswald, Juli 2010
























Dekan: Prof. Dr. Klaus Fesser………………………………………………

1. Gutachter:
Prof. Dr. Johannes F. Imhoff

2. Gutachter:
PD. Dr. Sabine Mundt

Tag der
Promotion:……16.09.2010……………………………………………………………






Hiermit erkläre ich, daß diese Arbeit bisher von mir weder an der
Mathematisch-Naturwissenschaftlichen Fakultät der Ernst-Moritz-Arndt-Universität
Greifswald noch einer anderen wissenschaftlichen Einrichtung zum Zwecke der
Promotion eingereicht wurde.

Ferner erkläre ich, daß ich diese Arbeit selbständig verfaßt und keine anderen
als die darin angegebenen Hilfsmittel benutzt habe.




Greifswald, den 19 Juli 2007 Unterschrift




Le Thi Anh Tuyet

a


.....................................................................................................1
1.1 Cyanobacteria.................................................................................................................. 1
1.1.1 Cyanobacterial physiology and morphology..................................................................... 1
1.1.2 Ecology of cyanobacteria................................................................................................ 2
1.1.3 Classification of cyanobacteria........................................................................................ 3
1.2 Cyanobacteria–a new and rich source of novel bioactive compounds with
pharmaceutical potential ........................................................................................................ 6
1.2.1 Antimicrobials.............................................................................................................. 9
1.2.2 Cytotoxic and antitumoural activities............................................................................. 12
1.2.3 Antiviral activity.......................................................................................................... 18
1.2.4 Toxins and other pharmacologically active compounds ................................................... 19
1.3 Aim of the work.............................................................................................................. 22


...........................................................................23
2.1 Biological materials....................................................................................................... 23
2.1.1 Soil cyanobacteria........................................................................................................ 23
2.1.2 Marine cyanobacteria ................................................................................................... 29
2.1.3 Bacteria, yeast, and cancer cell lines as test organisms .................................................... 31
2.2 Chemicals....................................................................................................................... 31
2.2.1 Cultivation of cyanobacteria ......................................................................................... 31
2.2.2 Cultivation of bacteria and yeast as test organisms.......................................................... 32
2.2.3 General laboratory chemicals.......................................................................................... 32
2.2.4 Chemical reagents........................................................................................................ 32
2.2.5 Fatty acid analysis ......................................................................................................... 33
2.3 Solvents............................................................................................................................ 33
2.4 Equipment in generally company, town, country ...................................................... 34
2.4.1 Cultivation of cyanobacteria ......................................................................................... 34
2.4.2 Extraction ..................................................................................................................... 34
2.4.3 Isolation of secondary metabolites................................................................................. 35
2.4.3.1 Thin layer chromatography (TLC).......................................................................... 35
2.4.3.2 Preparative TLC ................................................................................................... 35
2.4.3.3 Open column chromatography ............................................................................... 35
2.4.3.4 HPLC .................................................................................................................. 36
2.4.4 Agar plate diffusion test ................................................................................................. 36
2.4.5 Bioautographic TLC assay.............................................................................................. 37
2.4.6 Fatty acid analyses ....................................................................................................... 37
2. 5 Cultivation of cyanobacteria ....................................................................................... 38
2.5.1 The stock culture ......................................................................................................... 38
2.5.2 The batch culture ......................................................................................................... 38
2.5.3 The large scale culture.................................................................................................. 38
2.6 Extraction ....................................................................................................................... 39
2.6.1 Extraction of intracellular compounds............................................................................ 39
2.6.2 Extraction of extracellular compounds........................................................................... 40

2.7 Bioassays....................................................................................................................... 41
2.7.1 Assays for antimicrobial activity ................................................................................... 41
2.7.1.1 Agar diffusion assay.............................................................................................. 41
2.7.1.2 Bioautographic TLC assay..................................................................................... 42
2.7. 2 Assays for cytotoxic activity ........................................................................................ 43
2.8 Fractionation and isolation of the secondary metabolites of 6 cyanobacterial strains
................................................................................................................................................. 43
2.8.1 Fractionation and isolation of the secondary metabolites of Westiellopsis sp.VN................. 43
2.8.2 Fractionation and isolation of the secondary metabolites of Calothrix javanica ................. 45
2.8.3. Fractionation and isolation of the secondary metabolites of Scytonema ocellatum .............. 47
2.8.4 Fractionation and isolation of the secondary metabolites of Anabaena sp.......................... 49
2.8.5 Fractionation and isolation of the secondary metabolites of Nostoc sp. ............................. 50
2.8.6 Fractionation and isolation of the secondary metabolites of Lyngbya majuscula.................. 51
2.8.6.1 Method 1.............................................................................................................. 51
2.8.6.2 Method 2.............................................................................................................. 53
2.9 Structure elucidation of the isolated secondary metabolites................................... 55
2.9.1 Structure elucidation of compounds isolated from Westiellopsis sp.VN and Lyngbya majuscula
............................................................................................................................................. 55
2.9.2 Structure elucidation of compounds isolated from Calothrix javanica and Scytonema
ocellatum .........................................................